The present invention relates to compounds which are useful for treating pathological states which arise from or are exacerbated by angiogenesis, to pharmaceutical compositions comprising these compounds, and to methods of inhibiting angiogenesis in a mammal.
Angiogenesis, the process by which new blood vessels are formed, is essential for normal body activities including reproduction, development and wound repair. Although the process is not completely understood, it is believed to involve a complex interplay of molecules which regulate the growth of endothelial cells (the primary cells of capillary blood vessels). Under normal conditions, these molecules appear to maintain the microvasculature in a quiescent state (i.e. one of no capillary growth) for prolonged periods which may last for as long as weeks or, in some cases, decades. When necessary (such as during wound repair), these same cells can undergo rapid proliferation and turnover within a 5 day period (Folkman, J. and Shing, Y., The Journal of Biological Chemistry, 267(16), 10931-10934, (1992) and Folkman, J. and Klagsbrun, M., Science, 235, 442-447 (1987).
Although angiogenesis is a highly regulated process under normal conditions, many diseases (characterized as angiogenic diseases) are driven by persistent unregulated angiogenesis. Otherwise stated, unregulated angiogenesis may either cause a particular disease directly or exacerbate an existing pathological condition. For example, ocular neovascularization has been implicated as the most common cause of blindness and dominates approximately twenty eye diseases. In certain existing conditions, such as arthritis, newly formed capillary blood vessels invade the joints and destroy cartilage. In diabetes, new capillaries formed in the retina invade the vitreous, bleed, and cause blindness. Growth and metastasis of solid tumors are also dependent on angiogenesis (Folkman, J., Cancer Research, 46, 467-473 (1986), Folkman, J., Journal of the National Cancer Institute, 82, 4-6 (1989). It has been shown, for example, that tumors which enlarge to greater than 2 mm must obtain their own blood supply and do so by inducing the growth of new capillary blood vessels. Once these new blood vessels become embedded in the tumor, they provide a means for tumor cells to enter the circulation and metastasize to distant sites such as liver, lung or bone (Weidner, N., et al., The New England Journal of Medicine, 324(1), 1-8 (1991).
Several angiogenesis inhibitors are currently under development for use in treating angiogenic diseases (Gasparini, G. and Harris, A. L., J. Clin. Oncol., 13(3): 765-782, (1995), but there are disadvantages associated with these compounds. Suramin, for example, is a potent angiogenesis inhibitor but causes severe systemic toxicity in humans at doses required for antitumor activity. Compounds such as retinoids, interferons and antiestrogens are relatively safe for human use but have weak antiangiogenic effects. Irsogladine, an anti-tumor drug with low toxicity, has only weak anti-angiogenic effects. Thus there is still a need for compounds useful in treating angiogenic diseases in mammals.
In one embodiment of the present invention are disclosed compounds represented by Formula I 
or a pharmaceutically acceptable salt or prodrug thereof, where
L1 is selected from
(1) a covalent bond,
(2) xe2x80x94C(O)NR5(CH2)mxe2x80x94, where m is an integer from 0 to 4, and
R5 is selected from
(a) hydrogen
xe2x80x83and
(b) alkyl,
xe2x80x83and
(3) xe2x80x94N(R5)C(O)(CH2)mxe2x80x94, where (2) and (3) are drawn with their left ends attached to R1;
R1 is selected from
(1) alkyl,
(2) alkyl substituted with 1, 2, or 3 substituents selected from
(a) xe2x80x94NO2 
(b) xe2x80x94NR6R7 where R6 and R7 are independently selected from
(i) hydrogen,
(ii) alkyl,
(iii) arylalkyl,
(iv) an amino protecting group,
(v) alkanoyl, where the alkanoyl can be optionally substituted with xe2x80x94OR9,
(vi) (aryl)oyl,
(vii) alkoxycarbonyl,
xe2x80x83and
(viii) (heteroaryl)oyl,
xe2x80x83and
(c) alkoxycarbonyl,
(3) aryl substituted with 1, 2, 3, 4, or 5 substituents independently selected from
(a) xe2x80x94NR6R7,
(b) alkyl,
xe2x80x83and
(c) alkyl substituted with 1, 2, or 3 substituents selected from xe2x80x94NR6R7,
(4) xe2x80x94NR6R7,
xe2x80x83and
(5) xe2x80x94OR9;
R2 and R3 are selected from
(1) hydrogen
(2) xe2x80x94(CH2)nC(O)R8 where n is an integer from 0 to 4, and
R8 is selected from
(a) xe2x80x94OR9 where R9 is selected from
(i) hydrogen,
(ii) alkyl,
xe2x80x83and
(iii) alkyl substituted with 1 or 2 substituents selected from the group consisting of aryl
xe2x80x83and
(b) xe2x80x94NR5R10 where R5 is defined previously, and R10 is selected from
(i) hydrogen,
(ii) alkyl,
(iii) alkyl substituted with 1, 2, or 3 substituents independently selected from
(1xe2x80x2) xe2x80x94CO2R9 
xe2x80x83and
(2xe2x80x2) xe2x80x94C(O)NR6R7 
(iv) aryl,
xe2x80x83and
(v) arylalkyl, where (iv) and (v) can be optionally substituted with 1, 2, 3, 4, or 5 substituents independently selected from
(1xe2x80x2) alkyl,
(2xe2x80x2) alkanoyl,
(3xe2x80x2) xe2x80x94OR9,
(4xe2x80x2) xe2x80x94CO2R9,
(5xe2x80x2) alkanoyloxy,
(6xe2x80x2) carboxaldehyde,
(7xe2x80x2) cycloalkyl,
(8xe2x80x2) cycloalkenyl,
(9xe2x80x2) halo,
(10xe2x80x2) nitro,
(11xe2x80x2) perfluoroalkyl,
(12xe2x80x2) perfluoroalkoxy,
(13xe2x80x2) xe2x80x94NR6R7,
(14xe2x80x2) xe2x80x94SO2NR6R7,
(15xe2x80x2) xe2x80x94C(O)NR6R7,
(16xe2x80x2) aryloxy,
xe2x80x83and
(17xe2x80x2) aryl,
xe2x80x83and
(3) aryl, wherein the aryl is optionally substituted with 1, 2, or 3 substituents independently selected from
(a) xe2x80x94NR6R7 
xe2x80x83and
(b) xe2x80x94CO2R9,
provided that at least one of R2 and R3 is other than hydrogen;
R4 is selected from
(1) hydrogen,
(2) alkyl,
(3) cycloalkyl,
(4) xe2x80x94CO2R5,
(5) aryl,
xe2x80x83and
(6) aryl substituted with at least one of W, X, Y, or Z where W, X, Y, and Z are independently selected from
(a) alkyl,
(b) alkanoyl,
(c) xe2x80x94OR9,
(d) xe2x80x94CO2R9,
(e) alkanoyloxy,
(f) carboxaldehyde,
(g) cycloalkyl,
(h) cycloalkenyl,
(i) halo,
(j)nitro,
(k) perfluoroalkyl,
(l) perfluoroalkoxy,
(m) xe2x80x94NR6R7,
(n) xe2x80x94SO2NR6R7,
(o) xe2x80x94C(O)NR6R7,
(p) aryloxy,
xe2x80x83and
(q) aryl.
In another embodiment of the invention are disclosed methods of treating diseases comprising administering an effective amount of a compound having Formula I.
In yet another embodiment of the invention are disclosed pharmaceutical compositions containing compounds of Formula I.
Compounds of this invention include, but are not limited to,
N-[4-[N-(acetylglycyl)amino]benzoyl]-L-aspartic acid,
4-[[4-(aminomethyl)benzoyl]amino]-2-phenylbenzoic acid,
N-[4-[(7-amino-1-oxoheptyl)amino]benzoyl]-L-aspartic acid,
(S)-methyl 3-[[6-amino-2-[[(1,1-dimethylethoxy)carbonyl]amino]-1-oxohexyl]amino]-4-(1,1-dimethylethyl)benzoate,
(S)-methyl 3-[[2-(acetylamino)-6-amino-1-oxohexyl]amino]-4-(1,1-dimethylethyl)benzoate,
(S)-3-[[2-(acetylamino)-6-amino-1-oxohexyl)amino]-4-(1,1-dimethylethyl)benzoic acid,
(S)-methyl 4-[[2-[[(1,1-dimethylethoxy)carbonyl]amino]-6-[(phenylmethoxy)carbonyl]amino]-1-oxohexyl]amino]-2-[3-(phenylmethoxy)-phenyl]benzoate,
(S)-1,1-dimethylethyl 4-[[2-[[(1,1-dimethylethoxy)carbonyl]amino]-6-[(phenylmethoxy)carbonyl]amino]-1-oxohexyl]amino]-2-[3-(phenylmethoxy)phenyl]benzoate,
(R)-methyl 4-[[6-amino-2-[[(1,1-dimethylethoxy)carbonyl]amino]-1-oxohexyl]amino]-2-(3-hydroxyphenyl)benzoate,
(R)-methyl 4-[[6-amino-2-[[(1,1-dimethylethoxy)carbonyl]amino]-1-oxohexyl]amino]-2-(2-hydroxyphenyl)benzoate,
(S)-methyl 4-[[2-amino-6-[[(phenylmethoxy)carbonyl]amino]-1-oxohexyl]amino]-2-[(3-(phenylmethoxy)phenyl]benzoate,
(S)-methyl 4-[[2-(acetylamino)-6-[[(phenylmethoxy)carbonyl]amino]-1-oxohexyl]amino]-2-[(3-(phenylmethoxy)phenyl]benzoate,
(S)-1,1-dimethylethyl 4-[[6-amino-2-[[(1,1-dimethylethoxy)carbonyl]amino]-1-oxohexyl]amino-2-(3-hydroxyphenyl)benzoate,
(S)-methyl 4-[[2-(acetylamino)-6-amino-1-oxohexyl]amino]-2-(3-hydroxyphenyl)benzoate,
(S)-4-[[2-(acetylamino)-6-amino-1-oxohexyl)amino]-2-(3-hydroxyphenyl)benzoic acid,
(S)-N-[4-[[2-(acetylamino)-6-amino-1-oxohexyl]amino]-2-(3-hydroxyphenyl)benzoyl]-L-xcex1-asparagine,
(S)-N-[4-[[2-(acetylamino)-6-amino-1-oxohexyl]amino]-2-phenylbenzoyl]-L-xcex1-asparagine,
(S)-N-[4-[[2-(acetylamino)-6-amino-1-oxohexyl]amino]benzoyl]-L-xcex1-asparagine,
N-[(4-aminophenyl)acetyl]-L-aspartic acid, bis(1,1-dimethylethyl) ester,
(S)-N-[[4-[[2-amino-6-[[(phenylmethoxy)carbonyl]amino]-1-oxohexyl]amino]phenyl]acetyl]-L-aspartic acid,
(S)-N-[[4-[[2-(acetylamino)-6-[[(phenylmethoxy)carbonyl]amino]-1-oxohexyl]amino]phenyl]acetyl]-L-aspartic acid,
N-[2-[[4-[2-(acetylamino)-6-amino-1-oxohexyl]amino]phenyl]-1-oxoethyl]-L-aspartic acid,
(S)-N-[[4-[[6-amino-2-[[(1,1-dimethylethoxy)carbonyl]amino]-1-oxohexyl]amino]phenyl]acetyl]-L-aspartic aicd, bis(1,1-diemthylethyl) ester,
(S)-N-[[4-[(2,6-diamino-1-oxohexyl)amino]phenyl]acetyl]-L-aspartic acid,
(S)-ethyl 4-[[6-amino-2-[[(1,1-dimethylethoxy)carbonyl]amino]-1-oxohexyl]amino]benzeneacetate,
(S)-4-[[6-amino-2-[[(1,1-dimethylethoxy)carbonyl]amino]-1-oxohexyl]amino]benzeneacetic acid,
methyl 5-(((2S)-6-amino-2-((tert-butoxycarbonyl)amino)hexanoyl)amino)-4xe2x80x2-hydroxy(1,1xe2x80x2-biphenyl)-2-carboxylate,
(3S)-3-(((5-(((2S)-2-(acetylamino)-6-aminohexanoyl)amino)-4xe2x80x2-hydroxy(1,1xe2x80x2-biphenyl)-2-yl)carbonyl)amino)-4-amino-4-oxobutanoic acid,
methyl 3-(((2S)-6-amino-2-((tert-butoxycarbonyl)amino)hexanoyl)amino)-4-cyclohexylbenzoate,
tert-butyl (3S)-3-(((5-(((2S)-2-(acetylamino)-6-aminohexanoyl)amino)-3xe2x80x2-hydroxy(1,1xe2x80x2-biphenyl)-2-yl)carbonyl)amino)-4-amino-4-oxobutanoate,
5-(((2S)-6-amino-2-((tert-butoxycarbonyl)amino)hexanoyl)amino)-3xe2x80x2-hydroxy(1,1xe2x80x2-biphenyl)-2-carboxylic acid,
methyl 5-(((2S)-2,6-diaminohexanoyl)amino)-3xe2x80x2-hydroxy(1,1xe2x80x2-biphenyl)-2-carboxylate,
5-(((2S)-6-amino-2-((2,2-dimethylpropanoyl)amino)hexanoyl)amino)-3xe2x80x2-hydroxy(1,1xe2x80x2-biphenyl)-2-carboxylic acid,
methyl 5-(((2S)-6-amino-2-((2,2-dimethylpropanoyl)amino)hexanoyl)amino)-3xe2x80x2-hydroxy(1,1xe2x80x2-biphenyl)-2-carboxylate,
5-(((2S)-6-amino-2-(benzoylamino)hexanoyl)amino)-3xe2x80x2-hydroxy(1,1xe2x80x2-biphenyl)-2-carboxylic acid,
5-(((2S)-6-amino-2-((methoxycarbonyl)amino)hexanoyl)amino)-3xe2x80x2-hydroxy(1,1xe2x80x2-biphenyl)-2-carboxylic acid,
(4S)-4-((4-(((2S)-2-(acetylamino)-6-aminohexanoyl)amino)benzoyl)amino)-5-methylamino)-5-oxopentanoic acid,
4-(((2S)-6-amino-2-((tert-butoxycarbonyl)amino)hexanoyl)amino)benzoic acid,
(3S)-3-((4-(((2S)-2-(acetylamino)-6-aminohexanoyl)amino)benzoyl)amino)-4-amino-4-oxobutanoic acid,
methyl 4-(((2S)-6-amino-2-((tert-toxycarbonyl)amino)hexanoyl)amino)benzoate,
methyl 5-(((2S)-2-((tert-butoxycarbonyl)amino)hexanoyl)amino)-3xe2x80x2-hydroxy(1,1xe2x80x2-phenyl)-2-carboxylate,
4-(((2S)-6-amino-2-((tert-butoxycarbonyl)amino)hexanoyl)amino)-2-chlorobenzoic acid,
5-(((2S)-2-(acetylamino)-6-aminohexanoyl)amino)-N-(2-hydroxyphenyl)(1,1xe2x80x2-biphenyl)-2-carboxamide,
5-(((2S)-2-(acetylamino)-6-aminohexanoyl)amino)-N-(3-hydroxyphenyl)(1,1xe2x80x2-biphenyl)-2-carboxamide,
5-(((2S)-2-(acetylamino)-6-aminohexanoyl)amino)-N-(4-hydroxyphenyl)(1,1xe2x80x2-biphenyl)-2-carboxamide,
methyl 5-(((2S)-6-amino-2-(((benzyloxy)carbonyl)amino)hexanoyl)amino)(1,1xe2x80x2-biphenyl)-2-carboxylate,
5-(((2S)-2-((tert-butoxycarbonyl)amino)-6-((3-pyridinylcarbonyl)amino)hexanoyl)amino)-3xe2x80x2-hydroxy(1,1xe2x80x2-biphenyl)-2-carboxylic acid,
5-(((2S)-2-(acetylamino)-6-aminohexanoyl)amino)(1,1xe2x80x2-biphenyl)-2-carboxylic acid,
5-((6-aminohexanoyl)amino)-3xe2x80x2-hydroxy(1,1xe2x80x2-biphenyl)-2-carboxylic acid,
5-(((2S)-2-((tert-butoxycarbonyl)amino)hexanoyl)amino)-3xe2x80x2-hydroxy(1,1xe2x80x2-biphenyl)-2-carboxylic acid,
5-(((2S)-5-amino-2-((tert-butoxycarbonyl)amino)pentanoyl)amino)-3xe2x80x2-hydroxy(1,1xe2x80x2-biphenyl)-2-carboxylic acid,
(2S)-2-(((5-(((2S)-2-(acetylamino)-6-aminohexanoyl)amino)(1,1xe2x80x2-biphenyl)-2-yl)carbonyl)amino)butanedioic acid,
5-(((2S)-2-((tert-butoxycarbonyl)amino)-6-(methylamino)hexanoyl)amino)-3xe2x80x2-hydroxy(1,1xe2x80x2-biphenyl)-2-carboxylic acid,
5-(((2S)-2-(acetylamino)-6-aminohexanoyl)amino)-N-(4-(aminosulfonyl)phenethyl)(1,1xe2x80x2-biphenyl)-2-carboxamide,
ethyl 2-(((5-(((2S)-2-(acetylamino)-6-aminohexanoyl)amino)(1,1xe2x80x2-biphenyl)-2-yl)carbonyl)amino)benzoate,
ethyl 3-(((5-(((2S)-2-(acetylamino)-6-aminohexanoyl)amino)(1,1xe2x80x2-biphenyl)-2-yl)carbonyl)amino)benzoate,
ethyl 4-(((5-(((2S)-2-(acetylamino)-6-aminohexanoyl)amino)(1,1xe2x80x2-biphenyl)-2-yl)carbonyl)amino)benzoate,
5-(((2S)-2-(acetylamino)-6-aminohexanoyl)amino)-N-(4-(aminosulfonyl)benzyl)(1,1xe2x80x2-biphenyl)-2-carboxamide,
2-(((5-(((2S)-2-(acetylamino)-6-(((benzyloxy)carbonyl)amino)-hexanoyl)amino)(1,1xe2x80x2-biphenyl)-2-yl)carbonyl)amino)benzoic acid,
3-(((5-(((2S)-2-(acetylamino)-6-(((benzyloxy)carbonyl)amino)-hexanoyl)amino(1,1xe2x80x2-biphenyl)-2-yl)carbonyl)amino)benzoic acid,
4-(((5-(((2S)-2-(acetylamino)-6-(((benzyloxy)carbonyl)amino)-hexanoyl)amino)(1,1xe2x80x2-biphenyl)-2-yl)carbonyl)amino)benzoic acid,
2-(((5-(((2S)-2-(acetylamino)-6-aminohexanoyl)amino)(1,1xe2x80x2-biphenyl)-2-yl)carbonyl)amino)benzoic acid,
3-(((5-(((2S)-2-(acetylamino)-6-aminohexanoyl)amino)(1,1xe2x80x2-biphenyl)-2-yl)carbonyl)amino)benzoic acid, and
4-(((5-(((2S)-2-(acetylamino)-6-aminohexanoyl)amino)(1,1xe2x80x2-biphenyl)-2-yl)carbonyl)amino)benzoic acid.
The term xe2x80x9calkanoyl,xe2x80x9d as used herein, refers to an alkyl group attached to the parent molecular group through a carbonyl. The alkanoyl groups of this invention can be optionally substituted.
The term xe2x80x9calkoxy,xe2x80x9d as used herein, refers to an alkyl group attached to the parent molecular group through an oxygen atom. The alkanoyl groups of this invention can be optionally substituted.
The term xe2x80x9calkoxycarbonyl,xe2x80x9d as used herein, refers to an alkoxy group attached to the parent molecular group through a carbonyl. The alkoxycarbonyl groups of this invention can be optionally substituted.
The term xe2x80x9calkanoyloxy,xe2x80x9d as used herein, refers to an alkanoyl group attached to the parent molecular group through an oxygen atom. The alkanoyloxy groups of this invention can be optionally substituted.
The term xe2x80x9calkyl,xe2x80x9d as used herein, refers to a monovalent straight or branched chain group of one to twelve carbons derived from a saturated hydrocarbon by the removal of a hydrogen atom. The alkyl groups of this invention can be optionally substituted.
The term xe2x80x9camino,xe2x80x9d as used herein, refers to xe2x80x94NH2.
The term xe2x80x9caryl,xe2x80x9d as used herein, refers to a mono- or bicyclic carbocyclic ring system having one or two aromatic rings. The aryl group can also be fused to a cyclohexane, cyclohexene, cyclopentane or cyclopentene ring. The aryl groups of this invention can be optionally substituted.
The term xe2x80x9caryloxyxe2x80x9d as used herein, refers to an aryl group attached to the parent mplecular group through an oxygen atom. The aryloxy groups of this invention can be optionally substituted.
The term xe2x80x9carylalkyl,xe2x80x9d as used herein, refers to an aryl group attached to the parent molecular group through an alkyl group. The arylalkyl groups of this invention can be optionally substituted.
The term xe2x80x9ccarbonyl,xe2x80x9d as used herein, refers to xe2x80x94C(O)xe2x80x94.
The term xe2x80x9ccarboxaldehyde,xe2x80x9d as used herein, refers to xe2x80x94CHO.
The term xe2x80x9ccycloalkyl,xe2x80x9d as used herein, refers to a monovalent group of four to twelve carbons derived from a cyclic or bicyclic hydrocarbon having at least one carbon-carbon double bond. The cycloalkenyl groups of this invention can be optionally substituted.
The term xe2x80x9ccycloalkyl,xe2x80x9d as used herein, refers to a monovalent group of three to twelve carbons derived from a saturated cyclic or bicyclic hydrocarbon by the removal of a hydrogen atom. The cycloalkyl groups of this invention can be optionally substituted.
The term xe2x80x9chalo,xe2x80x9d as used herein, refers to xe2x80x94F, xe2x80x94Br, xe2x80x94Cl, or xe2x80x94I.
The term xe2x80x9cheteroaryl,xe2x80x9d as used herein, refers to an five- or six-membered aromatic ring containing at least one oxygen, nitrogen, or sulfur atom. The sulfur atoms can be optionally oxidized, and the nitrogen atons can be optionally oxidized or quaternized. Heterocycles of the invention are exemplified by those derived from furan, thiophene, pyrrole, imidazole, oxazole, thiazole, isoxazole, isothiazole, 1,2,3-oxadiazole, 1,2,3-triazole, 1,3,4-thiadiazole, pyridine, pyridazine, pyrirnidine, pyrazine, and 1,3,5-triazine. The heteroaryl groups of this invention can be optionally substituted.
The term xe2x80x9c(heteroaryl)oyl,xe2x80x9d as used herein, refers to a heteroaryl group attached to the parent molecular group theough a carbonyl.
The term xe2x80x9cN-protected aminoxe2x80x9d or xe2x80x9camino protecting group,xe2x80x9d as used herein, refers to groups intended to protect an amino group against undersirable reactions during synthetic procedures. Commonly used N-protecting groups are disclosed in Greene, T. W., and Wuts, P. G. M. (1991). Protectective Groups In Organic Synthesis (2nd ed.). New York: John Wiley and Sons. Preferred N-protecting groups are formyl, acetyl, benzoyl, pivaloyl, t-butylacetyl, phenylsulfonyl, benzyl, t-butyloxycarbonyl (Boc), and benzyloxycarbonyl (Cbz).
The term xe2x80x9cnitro,xe2x80x9d as used herein, refers to xe2x80x94NO2.
The term xe2x80x9cperfluoroalkoxy,xe2x80x9d as used herein, refers to a perfluoroalkyl group attached to the parent molecular group through an oxygen atom.
The term xe2x80x9cperfluoroalkyl,xe2x80x9d as used herein, refers to an alkyl group wherein all of the hydrogen atoms have been replaced with fluoride atoms.
The term xe2x80x9cpharmaceutically acceptable prodrugs,xe2x80x9d as used herein, presents those prodrugs of the compounds of the present invention which are, within the scope of sound medical judgement, suitable for use in contact with the tissues of humans and lower animals with undue toxicity, irritation, allergic response, and the like, commensurate with a reasonable benefit/risk ratio, and effective for their intended use, as well as the zwitterionic forms, where possible, of the compounds of the invention.
The term xe2x80x9cpharmaceutically acceptable salt,xe2x80x9d as used herein, represents those salts which are, within the scope of sound medical judgement, suitable for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio. Pharmaceutically acceptable salts are well known in the art. For example, S. M. Berge, et al. describe pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences, 1977, 66:1-19. The salts can be prepared in situ during the final isolation and purification of the compounds of the invention, or separately by reacting the free base function with a suitable organic acid. Representative acid addition salts include acetate, adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphersulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, fumarate, glucoheptonate, glycerophosphate, hemisulfate, heptonate, hexanoate, hydrobromide, hydrochloride, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palirtate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, toluenesulfonate, undecanoate, valerate salts, and the like. Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like, as well as nontoxic ammonium, quaternary ammonium, and amine cations, including, but not limited to ammonium, tetramethylammonium, tetraethylamrimonium, methylamine, dimethylamine, trimethylamine, triethylamine, ethylamine, and the like.
The term xe2x80x9cprodrug,xe2x80x9d as used herein, represents compounds which are rapidly transformed in vivo to the parent compound of the above formula, for example, by hydrolysis in blood. A thorough discussion is provided in T. Higuchi and V. Stella, Pro-drugs as Novel Delivery Systems, Vol. 14 of the A. C. S. Symposium Series, and in Edward B. Roche, ed., Bioreversible Carriers in Drug Design, American Pharmaceutical Association and Pergamon Press, 1987, both of which are incorporated herein by reference.
Compounds of the present invention can exist as stereoisomers where asymmetric or chiral centers are present. These compounds are designated by the symbols xe2x80x9cRxe2x80x9d or xe2x80x9cS,xe2x80x9d depending on the configuration of substitiuents around the chiral carbon atom. The present invention contemplates various stereoisomers and mixtures thereof. Stereoisomers include enantiomers and diastereomers, and equal mixtures of enantiomers are designated (xc2x1). Individual stereoisomers of compounds of the present invention can be prepared synthetically from commercially available starting materials which contain asymmetric or chiral centers or by preparation of racemic mixtures followed by resolution well-known to those of ordinary skill in the art. These methods of resolution are exemplified by (1) attachment of a mixture of enantiomers to a chiral auxiliary, separation of the resulting mixture of diastereomers by recrystallization or chromatography and liberation of the optically pure product from the auxiliary or (2) direct separation of the mixture of enantiomers on chiral chromatographic columns.
Endothelial Cell Migration Assay
The endothelial cell migration assay was performed essentially as described by Polverini, P. J. et al., Methods Enzymol, 198: 440-450 (1991). Briefly, Human Microvascular Endothelial Cells (HMVEC) were starved overnight in DMEM containing 0.1% bovine serum albumin (BSA). Cells were then harvested with trypsin and resuspended in DMEM with 0.1% BSA at a concentration of 1.5xc3x97106 cells/mL. Cells were added to the bottom of a 48-well modified Boyden chamber (Nucleopore Corporation, Cabin John, Md.). The chamber was assembled and inverted, and cells were allowed to attach for 2 hours at 37xc2x0 C. to polycarbonate chemotaxis membranes (5 xcexcm pore size) that had been soaked in 0.1% gelatin overnight and dried. The chamber was then reinverted and basic fibroblast growth factor (bFGF) and test substances were added to the wells of the upper chamber (to a total volume of 50 xcexcL); the apparatus was then incubated for 4 hours at 37xc2x0 C. Membranes were recovered, fixed and stained (DiffQuick, Fisher Scientific, Pittsburgh, Pa.) and the number of cells that had migrated to the upper chamber per 10 high power fields were counted. Background migration to DMEM+0.1% BSA was subtracted and the data reported as the number of cells migrated per 10 high power fields (400X) or when results from multiple experiments were combined, as the percent inhibition of migration compared to a positive control. The results are shown in Table 1.
The compounds of the invention, including but not limited to those specified in the example, possess anti-angiogenic activity. As angiogenesis inhibitors, such compounds are useful in the treatment of both primary and metastatic solid tumors and carcinomas of the breast; colon; rectum; oropharynx; hypopharynx; esophagus; stomach; pancreas; liver; gallbladder; bile; small intestine; urinary tract including kidney, bladder and urothelinum; female genital tract including cervix, uterus, ovaries, choriocarcinoma and gestational trophoblastic disease; male genital tract including prostate, seminal vesicles, testes and germ cell tumors; endocrine glands including thyroid, adrenal, and pituitary; skin including hemangiomas, melanomas, sarcomas, sarcomas arising from bone or soft tissues and Kaposi""s sarcomas; tumors of the brain, nerves, eyes, and meninges including astrocytomas, gliomas, gliblastomas, retinoblastomas, neuromas, neuroblastomas, Schwannomas and meningiomas; solid tumors arising from hematopoietic malignancies such as leukemias and including chloromas, plasmacytomas, plaques and tumors of mycosis fungoides and cutaneus T-cell lymphoma/leukemia; lymphomas including both Hodgkin""s and non-Hoggkin""s lymphomas; prophylaxis of autoimmune diseases including rheumatoid, immune and degenerative arthritis; ocular diseases including diabetic retinopathy, retinopathy of prematurity, corneal graft rejection, retrolental fibroplasia, neovascular glaucoma, rubeosis, retinal neovascularization due to macular degeneration and hypoxia; abnormal neovascularization conditions of the eye; skin diseases including psoriasis; blood vessel diseases including hemagiomas and capillary proliferation within atherosclerotic plaques; Osler-Webber Syndrome; myocardial angiogenesis; plaque neovascularization; telangiectasia; hemophiliac joints; angiofibroma; wound granulation; diseases characterized by excessive or abnormal stimulation of endothelial cells including intestinal adhesions, Crohn""s disease, atherosclerosis, scleroderma and hypertrophic scars (i.e. keloids) and diseases which have angiogenesis as a pathologic consequence including cat scratch disease (Rochele minalia quintosa) and ulcers (Helicobacter pylon). Another use is as a birth control agent which inhibits ovulation and establishment of the placenta.
The compounds of the present invention may also be useful for the prevention of metastases from the tumors described above either when used alone or in combination with radiotherapy and/or other chemotherapeutic treatments conventionally administered to patients for treating cancer. For example, when used in the treatment of solid tumors, compounds of the present invention may be administered with chemotherapeutic agents such as alpha inteferon, COMP (cyclophosphamide, vincristine, methotrexate and prednisone), etoposide, mBACOD (methortrexate, bleomycin, doxorubicin, cyclophosphamide, vincristine and dexamethasone), PRO-MACE/MOPP (prednisone, methotrexate (w/leucovin rescue), doxorubicin, cyclophosphamide, taxol, etoposide/mechlorethamine, vincristine, prednisone and procarbazine), vincristine, vinblastine, angioinhibins, TNP-470, pentosan polysulfate, platelet factor 4, angiostatin, LM-609, SU-101, CM-101, Techgalan, thalidomide, SP-PG and the like. Other chemotherapeutic agents include alkylating agents such as nitrogen mustards including mechloethamine, melphan, chlorambucil, cyclophosphamide and ifosfamide; nitrosoureas including carmustine, lomustine, semustine and streptozocin; alkyl sulfonates including busulfan; triazines including dacarbazine; ethyenimines including thiotepa and hexamethylmelamine; folic acid analogs including methotrexate; pyrimidine analogues including 5-fluorouracil, cytosine arabinoside; purine analogs including 6-mercaptopurine and 6-thioguanine; antitumor antibiotics including actinomycin D; the anthracyclines including doxorubicin, bleomycin, mitomycin C and methramycin; hormones and hormone antagonists including tamoxifen and cortiosteroids and miscellaneous agents including cisplatin and brequinar.
The compounds of the present invention may be used in the form of pharmaceutically acceptable salts derived from inorganic or organic acids. By xe2x80x9cpharmaceutically acceptable saltxe2x80x9d is meant those salts which are, within the scope of sound medical judgement, suitable for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, allergic response and the like and are commensurate with a reasonable benefit/risk ratio. Pharmaceutically acceptable salts are well-known in the art. For example, S. M. Berge, et al. describe pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences, 1977, 66: 1 et seq. The salts may be prepared in situ during the final isolation and purification of the compounds of the invention or separately by reacting a free base function with a suitable acid. Representative acid addition salts include, but are not limited to acetate, adipate, alginate, citrate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, camphorate, camphorsufonate, digluconate, glycerophosphate, hemisulfate, heptanoate, hexanoate, fumarate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethansulfonate (isethionate), lactate, maleate, methanesulfonate, nicotinate, 2-naphthalenesulfonate, oxalate, pamoate, pectinate, persulfate, 3-phenylpropionate, picrate, pivalate, propionate, succinate, tartrate, thiocyanate, phosphate, glutamate, bicarbonate, p-toluenesulfonate and undecanoate. Also, the basic nitrogen-containing groups can be quaternized with such agents as lower alkyl halides such asmethyl, ethyl, propyl, and butyl chlorides, bromides and iodides; dialkyl sulfates like dimethyl, diethyl, dibutyl and diamyl sulfates; long chain halides such as decyl, lauryl, myristyl and stearyl chlorides, bromides and iodides; arylalkyl halides like benzyl and phenethyl bromides and others. Water or oil-soluble or dispersible products are thereby obtained. Examples of acids which may be employed to form pharmaceutically acceptable acid addition salts include such inorganic acids as hydrochloric acid, hydrobromic acid, sulphuric acid and phosphoric acid and such organic acids as oxalic acid, maleic acid, succinic acid and citric acid.
Basic addition salts can be prepared in situ during the final isolation and purification of compounds of this invention by reacting a carboxylic acid-containing moiety with a suitable base such as the hydroxide, carbonate or bicarbonate of a pharmaceutically acceptable metal cation or with ammonia or an organic primary, secondary or tertiary amine. Pharmaceutically acceptable salts include, but are not limited to, cations based on alkali metals or alkaline earth metals such as lithium, sodium, potassium, calcium, magnesium and aluminum salts and the like and nontoxic quaternary ammonia and amine cations including ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, diethylamine, ethylamine and the like. Other representative organic amines useful for the formation of base addition salts include ethylenediamine, ethanolamine, diethanolamine, piperidine, piperazine and the like. Preferred salts of the compounds of the invention include phosphate, tris and acetate.
Compounds of this invention may be combined with pharmaceutically acceptable sustained-release matrices, such as biodegradable polymers, to form therapeutic pocompositions. A sustained-release matrix, as used herein, is a matrix made of materials, usually polymers, which are degradable by enzymatic or acid-base hydrolysis or by dissolution. Once inserted into the body, the matrix is acted upon by enzymes and body fluids. A sustained-release matrix is desirably chosen from biocompatible materials such as liposomes, polylactides (polylactic acid), polyglycolide (polymer of glycolic acid), polylactide co-glycolide (copolymers of lactic acid and glycolic acid) polyanhydrides, poly(ortho)esters, polypeptides, hyaluronic acid, collagen, chondroitin sulfate, carboxylic acids, fatty acids, phospholipids, polysaccharides, nucleic acids, polyamino acids, amino acids such as phenylalanine, tyrosine, isoleucine, polynucleotides, polyvinyl propylene, polyvinylpyrrolidone and silicone. A preferred biodegradable matrix is a matrix of one of either polylactide, polyglycolide, or polylactide co-glycolide (co-polymers of lactic acid and glycolic acid).
Compounds of this invention or combinations thereof may be combined with pharmaceutically acceptable excipients or carriers to form therapeutic compositions. A pharmaceutically acceptable carrier or excipient refers to a non-toxic solid, semi-solid or liquid filler, diluent, encapsulating material or formulation auxiliary of any type. The compositions may be administered parenterally, sublingually, intracisternally, intravaginally, intraperitoneally, rectally, bucally or topically (as by powder, ointment, drops, transdermal patch or iontophoresis device).
The term xe2x80x9cparenteral,xe2x80x9d as used herein, refers to modes of administration which include intravenous, intramuscular, intraperitoneal, intrasternal, subcutaneous and intraarticular injection and infusion. Pharmaceutical compositions for parenteral injection comprise pharmaceutically acceptable sterile aqueous or nonaqueous solutions, dispersions, suspensions or emulsions as well as sterile powders for reconstitution into sterile injectable solutions or dispersions just prior to use. Examples of suitable aqueous and nonaqueous carriers, diluents, solvents or vehicles include water, ethanol, polyols (such as glycerol, propylene glycol, polyethylene glycol and the like), carboxymethylcellulose and suitable mixtures thereof, vegetable oils (such as olive oil) and injectable organic esters such as ethyl oleate. Proper fluidity may be maintained, for example, by the use of coating materials such as lecithin, by the maintenance of the required particle size in the case of dispersions and by the use of surfactants. These compositions may also contain adjuvants such as preservatives, wetting agents, emulsifying agents and dispersing agents. Prevention of the action of microorganisms may be ensured by the inclusion of various antibacterial and antifungal agents such as paraben, chlorobutanol, phenol, sorbic acid and the like. It may also be desirable to include isotonic agents such as sugars, sodium chloride and the like. Prolonged absorption of the injectable pharmaceutical form may be brought about by the inclusion of agents, such as aluminum monostearate and gelatin, which delay absorption. Injectable depot forms are made by forming microencapsule matrices of the drug in biodegradable polymers such as polylactide-polyglycolide, poly(orthoesters) and poly(anhydrides). Depending upon the ratio of drug to polymer and the nature of the particular polymer employed, the rate of drug release can be controlled. Depot injectable formulations are also prepared by entrapping the drug in liposomes or microemulsions which are compatible with body tissues. The injectable formulations may be sterilized, for example, to by filtration through a bacterial-retaining filter or by incorporating sterilizing agents in the form of sterile solid compositions which can be dissolved or dispersed in sterile water or other sterile injectable media just prior to use.
Topical administration includes administration to the skin, mucosa and surfaces of the lung and eye. Compositions for topical administration, including those for inhalation, may be prepared as a dry powder which may be pressurized or non-pressurized. In non-pressurized powder compositions, the active ingredient in finely divided form may be used in admixture with a larger-sized pharmaceutically acceptable inert carrier comprising particles having a size, for example, of up to 100 micrometers in diameter. Suitable inert carriers include sugars such as lactose. Desirably, at least 95% by weight of the particles of the active ingredient have an effective particle size in the range of 0.01 to 10 micrometers. For topical administration to the eye, a compound of the invention is delivered in a pharmaceutically acceptable ophthalmic vehicle such that the compound is maintained in contact with the ocular surface for a sufficient time period to allow the compound to penetrate the corneal and internal regions of the eye, as, for example, the anterior chamber, posterior chamber, vitreous body, aqueous humor, vitreous humor, cornea, iris/cilary, lens, choroid/retina and sclera. The pharmaceutically acceptable ophthalmic vehicle may, for example, be an ointment, vegetable oil or an encapsulating material. Alternatively, a compound of the invention may be injected directly into the vitreous and aqueous humor.
The composition may be pressurized and contain a compressed gas such as nitrogen or a liquified gas propellant. The liquified propellant medium and indeed the total composition is preferably such that the active ingredient does not dissolve therein to any substantial extent. The pressurized composition may also contain a surface active agent such as a liquid or solid non-ionic surface active agent or may be a solid anionic surface active agent. It is preferred to use the solid anionic surface active agent in the form of a sodium salt.
Compositions for rectal or vaginal administration are preferably suppositories which may be prepared by mixing the compounds of this invention with suitable non-irritating excipients or carriers such as cocoa butter, polyethylene glycol or a suppository wax which are solids at room temperature but liquids at body temperature and therefore melt in the rectum or vaginal cavity and release the active compound.
Compounds of the present invention may also be administered in the form of liposomes. As is known in the art, liposomes are generally derived from phospholipids or other lipid substances. Liposomes are formed by mono- or multi-lamellar hydrated liquid crystals that are dispersed in an aqueous medium. Any non-toxic, physiologically acceptable and metabolizable lipid capable of forming liposomes can be used. The present compositions in liposome form may contain, in addition to a compound of the present invention, stabilizers, preservatives, excipients and the like. The preferred lipids are the phospholipids and the phosphatidyl cholines (lecithins), both natural and synthetic. Methods to form liposomes are known in the art. See, for example, Prescott, Ed., Methods in Cell Biology, Volume XIV, Academic Press, New York, N.Y. (1976), p. 33 et seq., which is hereby incorporated herein by reference.
When used in the above or other treatments, a therapeutically effective amount of one of the compounds of the present invention may be employed in pure form or, where such forms exist, in pharmaceutically acceptable salt form and with or without a pharmaceutically acceptable excipient. A xe2x80x9ctherapeutically effective amountxe2x80x9d of the compound of the invention means a sufficient amount of the compound to treat an angiogenic disease (for example, to limit tumor growth or to slow or block tumor metastasis) at a reasonable benefit/risk ratio applicable to any medical treatment. It will be understood, however, that the total daily usage of the compounds and compositions of the present invention will be decided by the attending physician within the scope of sound medical judgment. The specific therapeutically effective dose level for any particular patient will depend upon a variety of factors including the disorder being treated and the severity of the disorder; activity of the specific compound employed; the specific composition employed; the age, body weight, general health, sex and diet of the patient; the time of administration; the route of administration; the rate of excretion of the specific compound employed; the duration of the treatment; drugs used in combination or coincidental with the specific compound employed and like factors well known in the medical arts. For example, it is well within the skill of the art to start doses of the compound at levels lower than those required to achieve the desired therapeutic effect and to gradually increase the dosage until the desired effect is achieved. Total daily dose of compounds of this invention to be administered locally or systemically to a human or other mammal host in single or divided doses may be in amounts, for example, from 0.01 to 200 mg/kg body weight daily and more usually 1 to 300 mg/kg body weight. If desired, the effective daily dose may be divided into multiple doses for purposes of administration. Consequently, single dose compositions may contain such amounts or submultiples thereof to make up the daily dose.
It will be understood that agents which can be combined with the compound of the present invention for the inhibition, treatment or prophylaxis of angiogenic diseases are not limited to those listed above, but include, in principle, any agents useful for the treatment or prophylaxis of angiogenic diseases.
Abbreviations
Abbreviations which have been used in the descriptions of the scheme and the examples that follow are: NMM for 4-methylmorpholine; EDCI for 1-ethyl-3-[3-(dimethylamino)propyl]-carbodiimide hydrochloride; HOBT for hydroxybenztriazole; TFA for trifluoroacetic acid; THF for tetrahydrofuran; DMF for dimethylformamide.
Synthetic Methods
The compounds and processes of the present invention will be better understood in connection with the following synthetic schemes which illustrate the methods by which the compounds of the invention may be prepared. The compounds of this invention may be prepared by a variety of synthetic routes. A representative procedure is outlined in Scheme 1 where L1, R1, R2, R3 and R4 are defined previously unless indicated otherwise. Depending on the nature of L1, R1, R2, R3, and R4, protection and subsequent deprotection of other reactive groups can be required to successfully complete the described synthetic sequences. Commonly used protecting groups are disclosed in Greene, xe2x80x9cProtective Groups In Organic Synthesis,xe2x80x9d (John Wiley and Sons, New York (1981)), which is incorporated herein by reference. It will be readily apparent to one of ordinary skill in the art reviewing the synthetic route depicted below that other compounds within Formula I can be synthesized by the substitution of appropriate reactants and agents in the synthesis shown below. 
As exemplified in Scheme 1, a biaryl coupling was accomplished with methyl-2-chloro-4-nitrobenzoate and a boronic acid in the presence of a palladium catalyst such as tetrakis(triphenylphosphine)palladium(0). The nitro moiety was reduced to an amine with hydrogen gas in the presence of 10% palladium on carbon. Lysine was then coupled to the aniline derivative with EDCI, HOBt and a base preferably NMM. The ester was saponified with lithium hydroxide and aspartic acid coupled to the benzoic acid derivative with EDCI, HOBT and a base preferably NMM. The N-Boc group and t-butyl esters were removed with TFA to give the amino acid. The free amine was acetylated with acetyl chloride, and the CBZ group was removed under hydrogen gas in the presence of 10% palladium on carbon.